Tetrasodium 2,2''-diamino-8,8''-dihydroxy-4',4'''-{4-[bis(2-hydroxyethyl)amino]-1,3,5-triazine-2,6-diyldiimino}bis(naphthalene-1-azobenzene-2',6-disulfonate)

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

July 22nd to August 22nd, 2002

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2002

Report date:

2002

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method C.4-A (Determination of the "Ready" Biodegradability - Dissolved Organic Carbon (DOC) Die-Away Test)

Version / remarks:

92/69/EC

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 A (Ready Biodegradability: DOC Die Away Test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Red LF 6339

IUPAC Name:

Red LF 6339

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, industrial (adaptation not specified)

Details on inoculum:

The sludge was washed twice with tap water with centrifugation and the supernatant liquid phase was decanted. A homogenised aliquot of the final sludged suspension was weighed, thereafter dried and the ratio of wet to dry weight was calculated.
- Concentration of sludge: 4 g dry matter /lt; final concentration 30 mg dry material/lt
- Aeration: yes, during holding

Duration of test (contact time):

28 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

94.4 mg/L

Based on:

test mat.

Remarks:

corresponds to 30.2 mg/l DOC

Initial test substance concentration 2

Initial conc.:

94.3 mg/L

Based on:

test mat.

Remarks:

corresponds to 30.4 mg/l DOC

Details on study design:

TEST CONDITIONS
- Composition of medium: according to testing guideline
- Test temperature: 20-22 °C
- pH: measured in each test flask before the addition of the activated sludge inocolum and at the end of incubation.
- pH adjusted: yes. with a diluted sodium hydroxide solution.
- Aeration of dilution water: each flask was loosely covered with aluminium foil to allow exchange of air between the flask and the surrounding atmosphere.
- Continuous darkness: yes.
- Stirring: continuously by magnetic stirrers

TEST SYSTEM
- Culturing apparatus: 2000-ml Erlenmeyer flask, cleaned with alcoholic hydrochloric acid, rinsed with deionised water and dried. Filled with 1000 ml of test medium or test medium containing test item and/or reference item.
- Number of culture flasks/concentration: two for test item, two for procedure control, one for inoculum blank, abiotic control and toxicity control.
- Measuring equipment: DOC analysis-Shimadzu TOC-500 analyser.

SAMPLING
- Sampling frequency: day 0 (treatment day), 5, 7, 11, 14, 21 and 28 of the incubation period
- Sampling method: one sample of about 10 ml was taken from each flask per sampling date.
- Sample preparation: filtered through a 0.45 μm and analysed on the day of sampling.

CONTROL AND BLANK SYSTEM
- Inoculum blank: one replicate containing only activate sludge
- Abiotic sterile control: one replicate containing mercury dichloride. 94.0 mg/l corresponding to measured 31.7 mg/l DOC.
- Toxicity control: one replicate contating sodium benzoate (50 mg/l), test item (94.0 mg/l) and activated sludge. corresponds to 61.3 mg/l DOC.
- Procedure control: 10 ml aliquot of the stock solution and activated sludge. 50 mg/l reference item corresponding to measured 30.1 mg/l DOC and 31.2 mg/l DOC (2 replicates)

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

% Degradation

Parameter:

% degradation (DOC removal)

Value:

ca. 6

Sampling time:

28 d

Remarks on result:

other: mean value

Details on results:

Points of degradation plot (test substance):
1 % degradation after 5 d
3.5 % degradation after 7 d
5.5 % degradation after 11 d
3.5 % degradation after 14 d
1 % degradation after 21 d
5.5 % degradation after 28 d

Points of degradation plot (Toxicity control)
52 % after 14 days

BOD5 / COD results

Results with reference substance:

Points of degradation plot (reference substance):
100 % degradation after 5 d
102 % degradation after 7 d
103 % degradation after 11 d
99 % degradation after 21 d
100 % degradation after 28 d

Applicant's summary and conclusion

Validity criteria fulfilled:

not specified

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

biodegradation 6 % after 28 days

Executive summary:

The test item was investigated for its ready biodegradability in a "28-Day DOC Die-Away Test" according to EU Commission Directive 92/69 EEC, C.4-A (1992) and OECD Guideline for Testing of Chemicals, No. 301 A (1992). In the test flasks containing the test item and activated sludge (inoculum) the mean concentrations of dissolved organic the substance was found to be not biodegradable under the test conditions.

In the abiotic control, containing the test item and poisoned mineral medium, no degradation was noted after 28 days of exposure (based on DOC-measurements). The reference item sodium benzoate was completely biodegraded within 7 days of exposure, confirming the suitability of the activated sludge.In the toxicity control, containing the test item, the reference item sodium benzoate and activated sludge (inoculum), the initial DOC decreased by 52 % within 14 days of exposure. Thus, according to the test guidelines the test item can be assumed to be not inhibitory to activated sludge because degradation was >35 % within 14 days.